The present application proposes additional endpoints in the National Toxicology Program/Food and Drug Administration (NTP/FDA) 2-year bisphenol A (BPA) toxicity study with a specific focus on expanding prostate gland analysis. Prior research has shown that transient developmental exposure to low-dose BPA can enhance the carcinogenic susceptibility of the adult prostate gland to elevated estrogen levels upon aging. Identification of permanent changes in the prostate DNA methylome indicated that the molecular mechanisms of BPA reprogramming involve altered epigenetic memory. The goals of the proposed project are to expand the prostatic endpoints following chronic, oral BPA exposure to include analysis of periurethral prostatic ducts, epigenetic marks and stem cell reprogramming that will together add significant value to the GLP-compliant toxicology studies by providing a molecular framework to understand heightened disease susceptibility. The following specific aims are proposed: Aim 1: Expand the prostatic evaluation to include the periurethral prostatic ducts. Aim 2: Evaluate prostatic susceptibility to hormonal carcinogenesis in chronic BPA-exposed rats. Aim 3: Analyze DNA methylation and expression of estradiol (E2)/ BPA-reprogrammed genes in lateral prostates following chronic BPA exposure to identify molecular fingerprints of prostate reprogramming. Aim 4: Examine the stem/progenitor cells from BPA exposed prostates for self-renewal activity, differentiation potential and responsiveness to estradiol. Sprague-Dawley rats will be chronically treated with a range of BPA doses at the FDA animal facility and for Aim 2, treated with testosterone + estradiol (T+E) as adults for 1.5 years. Tissues shipped to the University of Illinois at Chicago (UIC) laboratory will be evaluated using histologic approaches, molecular analysis of DNA methylation and gene transcription, and prostate stem cell culture using a prostasphere assay. By examining carcinogenic susceptibility and identifying the molecular underpinnings of life-long prostate perturbations by prolonged BPA exposure, the present multidisciplinary approach will markedly enhance the weight-of-evidence assessment by the NTP/FDA using GLP-guideline studies.